Method for preventing peripheral nerve damage

ABSTRACT

A vitamin B 12  supplement composition comprising vitamin B 12  with and/or without added folic acid that is essentially free of antioxidants, such as vitamin C, as well as iron is disclosed. Also disclosed are methods of using this vitamin composition to prevent brain and nervous system damage, such as peripheral nerve damage, as well as pernicious anemia, such as where such anemia is caused by a deficiency of vitamin B 12  deficiency.

This application is a continuation-in-part of U.S. application Ser. No.09/291,372, filed Apr. 14, 1999, now abandoned which was a divisional ofU.S. application Ser. No. 08/544,330, filed Oct. 17, 1995, now U.S. Pat.No. 5,932,624, issued Aug. 3, 1999, the disclosures of which are herebyincorporated by reference in their entirety.

Homocystinuria is characterized by high serum homocysteine levels andleads to blood vessel damage, excretion of homocysteine in the urine,mental retardation, ectopia lentis, sparse blonde hair, convulsivetendency, thromboembolic episodes, and fatty changes of liver and isassociated with defective formation of cystathionine synthetase.

Homocysteine is a homolog of cysteine and is produced by thedemethylation of methionine, and is an intermediate in the biosynthesisof cysteine from methionine via cystathionine by cystathioninase.

High serum homocysteine-related blood vessel damage may account for upto 20% of U.S. heart attacks, 40% of strokes and 60% of peripheralvenous occlusions, in addition to those in the placenta associated withneural tube defects in about 2,000 infants a year.

It has recently been disclosed that the B vitamins, folic acid andvitamin B₁₂, by converting homocysteine to methionine, lower high serumhomocysteine and thereby protect against high serum homocysteine-relatedblood vessel damage and nerve damage. The major sources of folic acidare foods that are often not ingested in sufficient amount, namely freshfruits and vegetables, particularly the dark green leafy vegetables andorange juice. However, while vitamin B₁₂ is in all animal protein,including meat, fish, poultry, eggs, milk and milk products, there isnone in anything that grows out of the ground.

Folic acid and vitamin B₁₂ are members of the vitamin B complexnecessary for the normal production of red blood cells and nerve cells.Folic acid is present in peptide linkages in high quantities in liver,green vegetables and yeast. Vitamin B₁₂ is present in high quantities inliver and other animal products.

Many plant and animal tissues contain folic acid as reduced methyl orformyl polyglutamates. Folates act as co-enzymes for processes in whichthere is transfer of a 1-carbon unit, as in purine and pyrimidinenucleotide biosynthesis, amino acid conversions such as histidine toglutamic acid and generation and use of formate. Absorption takes placein the small intestine. In the gut epithelial cells, polyglutamates arereduced to dihydro- and tetrahydra-folates, and absorbed bound toprotein and transported in blood serum as methyl tetrahydrafolate. Someabsorbed folate is excreted in the bile and re-absorbed, together withan amount not absorbed and excreted in the stool.

Vitamin B₁₂ is necessary for taking a one-carbon unit from folic acidand delivering it to homocysteine to convert homocysteine to methionine.Vitamin B₁₂ and folic acid are necessary for normal nerve function aswell as for blood formation.

Vitamin B₆ is involved in a different pathway for getting rid of excesshomocysteine, which pathway is usually less important than theB₁₂-folate dependent pathway.

Vitamin supplements containing Folic Acid and/or vitamin B₁₂ and orVitamin B₆ are known, however, such supplements contain other vitamins,phytochemicals and minerals such as iron and copper, or otherantioxidant substances, including antioxidants, which destroy some ofVitamin B₁₂ and also some of the folic acid.

In accordance with an aspect of the present invention there is provideda multiple vitamin supplement composition comprising folic acid andvitamin B₁₂ that is essentially free of antioxidants.

In accordance with another aspect of the present invention there isprovided a multiple vitamin supplement composition comprising folicacid, vitamin B₁₂ and vitamin B₆ that is essentially free ofantioxidants.

In accordance with another aspect of the present invention there isprovided a method of administering a multiple vitamin supplementcomposition for lowering high serum homocysteine levels to protectagainst the incidence of heart attack and other blood vessel relateddisorders.

In accordance with another aspect of the present invention there isprovided a method of preparing a multiple vitamin supplement compositioncomprising folic acid and vitamin B₁₂ that is essentially free ofantioxidants.

In accordance with another aspect of the present invention there isprovided a method of preparing a multiple vitamin supplement compositioncomprising folic acid, vitamin B₁₂ and vitamin B₆ that is essentiallyfree of antioxidants.

In accordance with the primary aspect of the present invention there isprovided a multiple vitamin supplement composition comprising folic acidand vitamin B₁₂. The composition may also contain vitamin B₆, whereinthe composition is essentially free of antioxidants.

It has been shown that folic acid and vitamin B₁₂ each have the abilityto protect against high serum homocysteine-related blood vessel damage,as in some circumstances, does vitamin B₆.

One problem with previous attempts at using vitamin supplements toprevent such cardiovascular problems from developing is that folic acidsupplements taken alone are unsafe since they allow unrecognizedgenetically predisposed vitamin B₁₂ deficiency to produce irreversiblenerve damage in susceptible populations. Susceptible populations includethe elderly (those at least about 50 years of age) and women of AfricanAmerican descent in their child-bearing years.

Accordingly, the multiple vitamin supplement composition of the presentinvention comprises folic acid and vitamin B₁₂, and may also containvitamin B₆. The addition of vitamin B₁₂ lowers to normal the vitaminB₁₂-deficiency-produced high serum homocysteine found in millions of theelderly.

Vitamin B₆, the other B vitamin involved in homocysteine metabolism isalso added to the multiple vitamin supplement of the present invention.The addition of vitamin B₁₂ and vitamin B₆ further metabolizeshomocysteine and lowers serum homocysteine levels.

In a preferred embodiment, the multiple vitamin supplement of thepresent invention contains 400 micrograms of folic acid, 100 microgramsof vitamin B₁₂ and 10 milligrams of vitamin B₆.

The present invention departs from the prior art in the discovery thatthe multiple vitamin supplement composition as described above must beessentially free of antioxidants. Antioxidants, including but notlimited to other vitamins, minerals such as iron and copper, and otherphytochemicals, destroy not only the vitamin B₁₂ in the multiple vitaminsupplement but also some of the folic acid in the supplement once thevitamin dissolves in the alimentary tract.

By “essentially free” it is meant that the vitamin composition of thepresent invention must not contain an amount of antioxidants which wouldtend to damage and inactivate some of the vitamin B₁₂ and/or folic acidof the vitamin supplement when dissolved in the digestive tract. Thepresence of lower amounts of antioxidants would not render the vitamincomposition of the present invention ineffective or of reducedeffectiveness. Thus, supplemental vitamin C (a biochemically unbalancedsynthetic product), which drives free radical formation from iron andheme (Herbert et al, J. Nutr., 126 (suppl. 4):1213S-1220S (1996))destroys substantial vitamin B₁₂ by converting it to useless, or evenharmful, analogues. [Herbert, V., in Present Knowledge in Nutrition (7thEdition), Washington, D.C., ILSI Press (1996) pp. 191-205]. Free radicalgeneration also destroys substantial folate and gastric intrinsic factor[Shaw et al, Alcohol, 7:153-157 (1990)] Further, effective Jan. 1, 1998,the FDA has mandated that all grains also be fortified with 140 μg PGA(pteroylglutamic acid or folic acid) per 100 g grain. Such grains arecurrently fortified with iron due to an old mandate when negative ironbalance was more widespread than the current 6% of all Americans. Also,it has only been learned recently that about 12% of Americans could beharmed by fortifying grains with iron because these persons areheterozygous for hemochromatosis. Many cereals and other products arealso fortified with vitamin C.

More recently, based on Framingham data, Jacques et al (New England J.Medicine, 340:1449-1454 (1999)) have reported that, while fortificationof grains with folic acid (PGA) in elderly adults increased serum folateand decreased serum homocystaine, the difference in mean homocysteinewas largely due to differences in vitamin B₁₂ and vitamin B₆ statusbetween users of B vitamin supplements and non-users. This supportsearlier studies that by age 65, about 49% of otherwise healthy elderlyadults no longer absorb vitamin B₁₂ from food [see: Flynn et al, J. Am.Coll. Nutr., 16:258-267 (1997); Miller et al, FASEB J., 13:A936 (1997);Herbert, FASEB J., 13:A227 (1999)] as determined by low sodiumholotranscobalamin II (holo TC II), a surrogate Schilling test, and thatabout 60% of this 49% had vasculotoxically high (>17) serumhomocysteine, and that 100 μg of crystalline vitamin B₁₂ orally daily(with malabsorption, 500 μg of vitamin B₁₂ intranasally weekly)permanently sustains normal serum total and TCII vitamin B₁₂ levels andkeeps homocysteine normal. The reason why crystalline vitamin B₁₂ andnot vitamin B₁₂ from food is superior is that, in the naturalprogression of genetically predisposed gastric atrophy, loss of gastricacid and enzymes precedes loss of intrinsic factor (IF) by years. Thus,without the acids and enzymes, vitamin B₁₂ is not split from theproteins in food and is lost in the stool. However, crystalline vitaminB₁₂ is free of peptide bonds and is normally absorbed via thephysiologic IF-dependent mechanism. With no IF, about 1% of any oraldose of crystalline vitamin B₁₂ is absorbed by mass action diffusion.Only 0.1 μg vitamin B₁₂ need be absorbed daily to sustain otherwisenormal health. Thus, lower mean serum folate in the elderly is largevitamin B₁₂ deficiency gut megaloblastosis producing malabsorption offolate, corrected by vitamin B₁₂ therapy, and lower red cell folate issecondary both to availability of vitamin B₁₂ to transport folate intored cells and to keep it in the red cells by polyglutamating it.

The components of the multiple vitamin supplement of the presentinvention are co-enzymes which act in accordance with methioninesynthetase and cystathioninase and facilitate the production ofmethionine and cysteine from homocysteine. This lowers the serum levelof homocysteine and the high level of heart attacks and other vasculardamage and nerve damage associated therewith. The presence ofantioxidants in the vitamin composition tends to inactivate the vitaminB₁₂ and folic acid components of the vitamin composition once thevitamin composition is processed in the alimentary tract.

Therapeutic treatment with the multiple vitamin supplement of thepresent invention may involve administration to personsprophylactically, that is to prevent, retard or reduce the severity offuture occurrence of the disease or its clinical manifestations.

In accordance with the foregoing, the present invention relates to aprocess of preventing vitamin B₁₂-deficiency induced brain and nervoussystem damage in a human at least 50 years of age comprisingadministering to said human a composition comprising at least about 25μg, or a range of at least about 25 μg to about 1000 μg of vitamin B₁₂wherein said vitamin B₁₂ is substantially free of antioxidants. In aspecific embodiment, such composition comprises about 100 μg of vitaminB₁₂ which could be plus or minus 10 μg.

Among the antioxidants especially to be avoided in such compositions isvitamin C, whether naturally present or added. Obviously, noantioxidants of any kind should be added to any of the compositionsdisclosed herein (although such antioxidants may be present during thepreparation of such vitamins provided that they are removed afterward,either completely or at least to a level where they have virtually noeffect on the vitamin components of the present invention). Also to beavoided is the presence of iron (Fe) in such compositions.

In a separate embodiment, the present invention finds especiallyadvantageous use in situations where said human exhibits gastricatrophy.

In accordance with the present invention, the processes disclosed hereinare especially useful wherein said brain and nervous system damagecomprises brain cell and/or peripheral nerve damage.

Because such antioxidants may be present in vitamin preparations usefulin forming the compositions of the present invention, the presentinvention also relates to processes as already described but whereinsaid vitamin B₁₂ has been tested for the presence of antioxidant andshown to be free of antioxidant. Such testing is commonly performed byliquifying a sample of the product (i.s., the vitamin, or vitamincomposition, or vitamin supplement) to be tested in a solution atstomach pH, and another sample at neutral pH, incubating for 30 minutes(the gastric half-emptying time) anf then assaying the amount ofvitamin, or vitamins, remaining as compared to the amount prior toincubation.

The present invention also relates to processes such as those describedherein where the composition also comprises folic acid in levelsdisclosed herein.

Pernicious anemia is, in general, a vitamin B₁₂ deficiency disease thatconstitutes a form of homocysteine vasculotoxicity and neurotoxicity.

The present invention further relates to a process of preventingpernicious anemia in a human comprising administering to said human acomposition comprising at least about 25 μg, or a range of at leastabout 25 μg to about 1000 μg of vitamin B₁₂ wherein said vitamin B₁₂ issubstantially free of antioxidants. In a specific embodiment, suchcomposition comprises about 100 μg of vitamin B₁₂, which could be plusor minus 10 μg.

Among the antioxidants especially to be avoided in such compositions forpreventing pernicious anemia is added vitamin C. Obviously, noantioxidants of any kind should be added to any of the compositionsdisclosed herein (although such antioxidants may be present during thepreparation of such vitamins provided that they are removed afterward,either completely or at least to a level where they have virtually noeffect on the vitamin components of the present invention). Also to beavoided is the presence of iron (Fe) in such compositions.

In a separate embodiment, the present invention for preventingpernicious anemia finds especially advantageous use in situations wheresaid human exhibits gastric atrophy. Thus, regardless of whether thevitamin composition, or supplement, or fortificant, of the presentinvention is to be utilized to prevent. pernicious anemia, or peripheralnerve damage, or other condition, 1% of any oral dose of vitamin B₁₂ ifabsorbed by mass action diffusion even after gastric atrophy, whichcommonly begins in humans as early as age fifty and becomes increasingmore prevalent at later ages. In fact, every person between age 50 andage 100 experiences some form of gastric atrophy in a geneticallypredetermined way. Such atrophy destroys the ability to absorb all ofthe small doses of vitamin B₁₂ (about 1 to 5 μg) eaten in daily meat,fish, poultry, and other animal products (since there is no vitamin B₁₂in any vegetable matter).

It has also been known for some time that the minimal daily absorbedrequirement of vitamin B₁₂ to sustain normal health is only about 0.1μg. Consequently, even with gastric atrophy and an inability to absorbthe vitamin B₁₂ in food, a pill containing at least about 25 μg ofvitamin B₁₂ daily permits absorption of about 0.25 μg (1%)±50% (or 0.125μg) thereof, thus guaranteeing sufficient absorbed vitamin B₁₂ eventhough food vitamin B₁₂ may be totally unabsorbable.

One problem of gastric atrophy is that as it progresses one losesgastric secretion of enzymes and acid which split vitamin B₁₂ from itspeptide bonds in food so that no food vitamin B₁₂ is absorbed. Thus, theaforementioned 1% absorbability by mass action diffusion applies only tocrystalline vitamin B₁₂ and not to vitamin B₁₂ in food (because thelatter is all protein bound). Because of this protein-binding, thecompositions disclosed herein should preferably not be taken within anhour (before or after) of eating (since gastric emptying halftime isabout 30 minutes).

It is also important to note that the compositions disclosed accordingto the present invention can be included as supplements or fortificantsin foods, such as in breads, cerals, and postas. The levels of vitaminB₁₂ disclosed herein should provide no problem with such uses. Forexample, use of 1000 μg of vitamin B₁₂ in a loaf of bread might tend tocause a pink coloration thereby discouraging sale and consumptionwhereas use of only 25 μg to about 500 μg of vitamin B₁₂ in a loaf ofbread is sufficient to provide the needed dose but does not cause anyunwanted coloration of the bread.

Because antioxidants may be present in vitamin preparations useful informing the compositions for preventing pernicious anemia, the presentinvention also relates to processes as already described but whereinsaid vitamin B₁₂ has been tested for the presence of antioxidant andshown to be free of antioxidant.

The compositions useful in the processes of the present invention forpreventing pernicious anemia may also comprise folic acid in levelsdisclosed below. Such compositions may also be in the form of a smartpill.

The multiple vitamin supplement composition of the present inventioncontains a therapeutically effective amount of folic acid, vitamin B₁₂and may or may not also contain vitamin B₆ with the composition beingessentially free of antioxidants. The vitamin composition may beadministered with a pharmaceutically acceptable carrier. Apharmaceutically acceptable carrier may be any compatible, non-toxic,non-antioxidant substance suitable to deliver the components. Thesupplement may contain other pharmaceutically acceptable substances asrequired to approximate physiological conditions such as a pH adjustingand buffering agent, disbursing agents, toxicity adjusting agents,flavoring agents and like. The concentration of the components in theseformulations may vary and will be selected primarily on the particulardosage and mode of administration selected. Methods for preparingsupplements are well-known or will be apparent to those skilled in theart and are described in more detail in, for example, Remington'sPharmaceutical Science, 15th ed., Mack Publishing Company, Easton, Pa.

The supplement is useful for oral administration. The supplement may beformulated in a variety of dosage forms, such as tablets, capsules, oralsolutions or suspensions.

Preferably, the supplement is administered orally. For oraladministration, solid or fluid dosage forms can be prepared. Forpreparing solid compositions such as tablets, the components are mixedwith conventional ingredients, such as talc, magnesium stearate, andfunctionally similar materials, as pharmaceutical carriers. Capsules areprepared by mixing the components with an inert pharmaceutical diluentand filling the mixture into a hard gelatin capsule. Soft gelatincapsules are prepared by machine encapsulation of a slurry of thecomponents with an acceptable vegetable oil, light liquid petrolatum orother inert oil. Fluid unit dosage forms for oil administration such asserum and suspensions can be prepared. The components may be dissolvedin an aqueous vehicle together with sugar, sweetening and flavoringagents and preservatives to form a serum. Suspensions can be preparedwith an aqueous vehicle and a disbursing agent such as acacia,tragacanth, methylcellulose and the like. In accordance with theinvention, any carrier, filler or other substance associated with thecomponents of the invention used to prepare a tablet, capsule or thelike must be essentially free of anti-oxidants.

In an alternate embodiment of the present invention, separate vitamincompositions may be prepared in accordance with the invention by themethods described above with each containing only Folic acid or VitaminB₁₂. These tablets are essentially free of anti-oxidant substances. Inthis manner, one or the other component of Folic acid or Vitamin B₁₂ canbe taken alone such that a user, or their physician, may have morecontrol over the quantity of intake of Folic Acid or Vitamin B₁₂,without be forced to also alter the level of intake of the other.

The compositions containing the multiple vitamin supplement componentsmay be administered for the prevention or therapeutic treatment of highserum homocysteine disorders, including prophylactic treatment. Intreatment of patients diagnosed with high serum homocysteine levels, thesupplement may be administered to a person in an amount sufficient toreduce serum homocysteine levels to normal. In prophylactic treatment,the supplement may be administered to a person who may be at risk ofhaving a high serum homocysteine blood vessel or nerve related disorder,but has not been diagnosed as having such an disorder. An amountadequate to accomplish any of these effects is referred to as a“therapeutically effective” amount. Unit dosages effective for this usewill depend upon the severity of the disorder and the general state ofthe person's health, but will generally range from 25 to about 400micrograms of folic acid, 25 to about 1,000 micrograms of vitamin B₁₂,and 5 to about 20 milligrams of vitamin B₆, with 400 micrograms of folicacid, 100 micrograms of vitamin B₁₂ and 10 milligrams of vitamin B₆being preferred. The multiple vitamin supplement may be administered indaily dosages and over a period of time with a frequency and durationsufficient to yield a “therapeutically effective” amount, i.e., anamount sufficient to reduce serum homocysteine levels to normal.

What is claimed:
 1. A process of preventing vitamin B₁₂-deficiencyinduced brain and nervous system damage in a human at risk thereofcomprising administering to said human a composition comprising at leastabout 25 μg of vitamin B₁₂ wherein said vitamin B₁₂ is substantiallyfree of antioxidants.
 2. The process of claim 1 wherein said compositioncomprises at least about 25 μg to about 1000 μg of vitamin B₁₂.
 3. Theprocess of claim 1 wherein said composition comprises about 100 μg ofvitamin B₁₂.
 4. The process of claim 1 wherein said human is at leastabout 50 years of age.
 5. The process of claim 1 wherein said humanexhibits gastric atrophy.
 6. The process of claim 1 wherein said brainand nervous system damage is selected from the group consisting of braincell damage and peripheral nerve damage.
 7. The process of claim 1wherein said is free of vitamin C and iron (Fe).
 8. The process of claim1 wherein said vitamin B₁₂ has been tested for the presence ofantioxidant and shown to be free of antioxidant.
 9. The process of claim1 wherein said composition also comprises folic acid.